The invention generally relates to improved, automated liquid handling methods applicable to filtration and preparative chromatography of liquids for use in the pharmaceutical and biotechnology industries. With the invention, a variety of separation techniques are handled in a yield-enhancing automated manner. The invention enhances separation processes such as tangential flow filtration (TFF) (also called cross flow filtration), direct flow filtration (DFF) (also called dead end filtration or normal flow filtration), as well as preparative chromatography applications.
In the pharmaceutical and biotechnology industries, the use of preparative chromatograpy, direct flow filtration (DFF) and tangential flow filtration (TFF), including micro-,ultra-, nano-filtration and diafiltration are well-established methods for the separation of dissolved molecules and/or suspended particulates.
In direct flow filtration (DFF), a filtration device is used that has one inlet and one outlet. The total (100%) solution volume is forced through a porous filter. DFF devices are typically single-use devices. Such membrane filters or depth filters are commercially available in different filter area sizes as well as different pore sizes. Depending upon the selected pore size, molecules or particulates smaller than the average membrane pore size will pass (together with solvent) through the filter. Thus, direct flow filtration (DFF) devices allow for the selective removal of particulates, bacteria, viruses, cell debris and large macro molecules.
In contrast, tangential flow filtration (TFF) devices have one inlet, one retentate outlet and at least one permeate outlet. In TFF, the retentate is repeatedly re-circulated with the objective of improving filtration efficiency and enhancing the permeate yield. The re-circulated retentate solution pathway runs parallel to the membrane surface and is pumped past the membrane with sufficient velocity to ensure a surface cleaning action. Furthermore, a sufficiently high trans-membrane pressure (TMP) or differential pressure (DP) is applied across the filtration device to ensure an optimal permeate flux across the membrane. However, only a relatively small amount of permeate is collected during each retentate volume-pass, and thus a significant processing time is typically associated with TFF procedures.
For optimal results, both DFF and TFF demand careful attention to filter porosity, filter area as well as required differential pressures and selected pump rates. However, filtration devices tend to clog when used over an extended period of time and must be timely replaced. Clogging of a filtration device occurs: (1) when the membrane pores become obstructed, typically with trapped cells, particulate matter, cell debris or the like or (2) when the feed channel (into a TFF device) becomes obstructed by solids or colloidal material and/or cell debris. This clogging of the feed channel or membrane pores results in a decreased liquid flow across the porous filter membrane. The result is a change in system pressure which, if not properly addressed, runs the risk of serious detriment to the operation which incorporates the filtration procedure.
Attempts to address these concerns and difficulties have included the development and use of semi-automated filtration systems. These types of systems utilized either manually controlled recirculation pumps or pumps which are controlled by a timing device which will stop pump action after a preset filtration time has elapsed. It is also typical to monitor line pressure through the use of an analog or a digital pressure gauge, usually located between the pump and the filter device. When the gauge reads a certain line pressure level, typically one specified by the manufacturer of the filter device, the filtration must be stopped and the old filter must be replaced with a new one. At times, it is not possible to accurately predict the time at which the pumping action must be stopped in order to avoid overtaxing the filter device. Accordingly, prior art systems which rely solely on timing are not entirely satisfactory.
Prior art filtration technology such as that referred to above also is disadvantageous because it is typically very labor intensive. This prior technology also has additional, serious shortcomings for safe and efficient operation. One shortcoming is that the filtrate yield frequently is not quantitative because of unpredictable solution particulate loads. Thus, for a given re-circulation volume and pump rate, the filtrate yield may differ from case to case, depending upon the amount of pore-sized particulate suspended in the recirculation solution. Another shortcoming is a direct result of back pressure build up due to clogging and gel layer formation. Rapid back pressure build up at times causes bursting of the filter membrane and/or the filter housing, resulting in costly spillage and/or filtrate contamination. Excessive filter inlet pressure also frequently leads to blow-off of tube connections such as at the filter inlet, resulting in costly spillage of retentate, for example. Because of these types of shortcomings, manual and semi-automated filtration systems need to be constantly monitored, which greatly contributes to the high labor intensity of such approaches.
With specific reference to TFF, the use of TFF for concentrating bio-molecules such as dilute protein solutions has a number of challenges associated with same that are related to solution viscosity changes during the TFF process. As the TFF operation progresses, solvent (typically water and small, undesirable buffer species) are removed from the dilute starting material and are collected as permeate. Under normal conditions, the selected porosity of the TFF device prevents the protein molecules from crossing the filter membrane. The progressive removal of solvent gives rise to a steady increase in the retentate (e.g. protein) concentration accompanied by a steady increase in retentate viscosity. The concentration and viscosity changes cause a general increase in the TFF system pressure, which complicates efforts at safe and optimal completion of the TFF operation.
It would be beneficial to achieve safe and optimal TFF operation, which implies a TFF procedure that maximizes permeate collection in the shortest time, within safe operating parameters, without generating excessive wall concentrations of product (e.g. protein) at the membrane surface, all while taking into account shortcomings of TFF components such as pumps, valves and the like. For example, selection of the TFF pump may be important in reducing destruction of retentate due to pump shear and or heat denaturation. These considerations come into play in attempting to achieve safe and optimum TFF operation. Typically, current TFF practices require frequent, manual adjustment of system pressured and/or flow rates over many hours in attempting to achieve safe and optimal TFF operations.
Filtration arrangements as described in Schick U.S. Pat. No. 5,947,689, incorporated hereinto by reference, provide for quantitative capability with TMP pressure monitoring. Such a filtration approach allows for rapid and safe filtration without concern of losing product, particularly pharmaceutical products or biotechnology products which can be extremely expensive, difficult to replace, and can represent the investment of many hours of prior processing. This patent describes coaxing the maximum life out of a filtration device without running the risk of generating operational conditions which can lead to excessive back pressure build up near the end of the life of the filtration device.
Filtration arrangements also are described in Schick U.S. Pat. No. 6,350,382 for effecting at least safe and automated TFF operational capabilities. Schick U.S. Pat. No. 6,607,669 describes effecting at least safe and automated TFF operational capabilities including automated diafiltration capabilities. Each of these patents and other references noted herein are incorporated by reference hereinto.
Filtration (DFF) rates which are too fast cause premature filter plugging or filter failure and related problems. Because of this, operators of filtration and column loading units have been known to set the equipment at an initially low filtration rate or flow rate and gradually increase the rate. Alternatively, the units are set up to provide a very large filtration surface area in order to thereby reduce the pump loading at the filter. Other current solutions include setting a low pump rate and maintaining same low throughout the loading cycle, which of course results in a lengthy load time. By another approach, a higher initial pump rate is selected, the filtering progress is monitored, and the pump rate is manually decreased, leading to a very uneconomic situation.
Previous liquid handling methods for direct flow filtration, tangential flow filtration and/or chromatographic column loading often have proceeded according to either a constant low pump rate or a constant safe pump pressure. These are time-consuming and typically require operator monitoring and/or intervention.
Accordingly, there is a need for improvements in liquid handling for valuable and vulnerable pharmaceutical and biotechnology compositions, components and products. There is also a need for such liquid handling to proceed with minimal operator monitoring requirements while providing safe, automated and optimal separation capabilities.